This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Our long-term goal is to establish a cell coculture model with a defined microenvironment to systematically study the electromechanical coupling of adult stem cells with cardiac myocytes at the single-cell level. The objective for this project is to understand how bone marrow stem cells electrically couple with cardiac myocytes and how stem cell differentiation will affect this coupling. We will investigate the electrical coupling of bone marrow stem cells in an engineered cardiac myocyte-coculture model. In this model, identical cell-culture microwells will be fabricated to allow creation of a defined stem cell-myocyte interface in each well through application of a laser beam. To test the hypothesis that the electrical coupling between stem cells and myocytes will be regulated by the spatial arrangements of cells and the temporal stages of stem cell differentiation, the following specific aims will be addressed: * To determine temporal and spatial extent of the electrical coupling between stem cells and cocultured myocytes at the cellular level * To identify effects of different connexins in stem cell-myocyte electrical coupling * To determine teffects of stem cell differentiation on stem cells'electrical coupling with cardiac myocytes Heart disease is America's leading cause of death for both men and women, accounting for nearly 40% of all annual deaths. By adding to the current knowledge of electrical coupling between bone marrow stem cells and heart muscle cells, our research will contribute to the recent effort in regenerative medicine to restore the function of a damaged heart using the patient's own cells. The significance of this research is that it provides an experimental model by which various stem cells are subjected to a highly controlled microenvironment during their functional differentiation. With application of the laser technique in this model, exact temporal and spatial constraints on electrical coupling and integration between stem cells and myocytes can be investigated. The results of this research will significantly advance our fundamental knowledge of the functional integration between stem cells and myocytes. In addition to 50% release time, Clemson will provide more than 2,500 sq ft of lab space and unlimited access to all Core facilities, which are staffed and provide with state-of-the-art equipment.